Determination of genetic stability in long-term somatic embryogenic cultures and derived plantlets of cork oak using microsatellite markers.

Microsatellites were used to test genetic stability in somatic embryos (SE) of Quercus suber L. The SE were obtained by a simple somatic embryogenesis protocol: leaf explants from two adult plants (QsG0, QsG5) and from two juvenile plants (QsGM1, QsGM2) were inoculated on Murashige and Skoog (MS) medium with 2,4-dichlorophenoxyacetic acid and zeatin. Calluses with primary embryogenic structures were transferred to MSWH (MS medium without growth regulators) and SE proliferated by secondary somatic embryogenesis. High morphological heterogeneity was found among cotyledonary SE. However, converted plants looked morphologically normal with well-developed rooting systems and shoots. The genetic stability of the plant material during the somatic embryogenesis process was evaluated by using six to eight nuclear microsatellites transferred from Q. myrsinifolia Blume, Q. petraea (Matts.) Liebl. and Q. robur L. Five of eight microsatellites distinguished among the genotypes analyzed, and for QsG0, QsGM1 and QsGM2, uniform microsatellite patterns were generally observed within and between SE and the respective donor genotypes. For genotype QsG5, the same pattern was observed in all samples analyzed except one, where the mutation percentage was 2.5%. We conclude that microsatellite markers can be used to assess genetic stability of clonal materials and to determine genetic stability throughout the process of somatic embryogenesis. The simple somatic embryogenesis protocol described has potential for the commercial propagation of Q. suber because it results in a low percentage of mutations.